Engineering synthetic biology routes for protein analysis using microfluidic systems

Student thesis: Doctoral Thesis

Abstract

Microfluidics is increasingly being used in many areas of biotechnology and chemistry to achieve reduced reagent volumes, improved performance, integration, and parallelism, among other advantages. The sub-millimetre dimensions of microchannels tend to reduce reagent consumption and waste production, consequently leading to cost savings and permitting precious samples to be relished and divided up for additional assays. Furthermore, the system allows for parallelisation, this way many identical reactions or assays can be replicated on a single microfluidic chip hence increasing throughput, a quality very much desired by pharmaceutical companies.Different steps in a complex process can be combined into a single chip to enhance ease of use, portability and reducing human error—for example, in medical diagnostic devices.Membrane proteins are of great importance as subjects of physiological study,drug targets in high-throughput assays for drug discovery and drug safety screening1. Microfluidic devices can be used for manipulating and analysing proteins, greatly benefiting many of the applications mentioned. The ability to produce artificial membranes within a microfluidic platform is crucial for the realization of these advantages. Current methods of producing and studying artificial cell membranes are typically low-throughput and not automated.This thesis presents the development of a fully integrated microfluidic system for the production of artificial lipid bilayers based on the miniaturisation of dropletinterface-bilayer (DIB) techniques. The platform uses a microfluidic design that enables formation, alternation, controlled positioning and long-term storage of arrays of droplet-interface-bilayers (DIBs) to mimic cell membrane processes. By encapsulating the desired cocktail of liposomes and metabolites into phospholipid stabilized water-in-oil (W/O) droplets, hundreds of DIBs were characterized. To ensure robustness of operation, we have investigated how lipid concentration,immiscible phase flow velocities and the device geometrical parameters affect the system performance. Finally, proof-of-concept data is shown where diffusive transport of molecules and ions across on-chip DIBs can be studied and quantified using fluorescence-based assays. The ability to quantitatively identify DIB permeation values demonstrates the suitability of our system for investigating processes occurringacross an artificial lipid bilayer in a miniaturised and scalable format.
Date of Award25 Sep 2019
Original languageEnglish
Awarding Institution
  • University Of Strathclyde
SponsorsEPSRC (Engineering and Physical Sciences Research Council)
SupervisorMichele Zagnoni (Supervisor) & Deepak Uttamchandani (Supervisor)

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